This proposal seeks to develop engineered intracellular antibodies (intrabodies) as novel potential clinical reagents and drug discovery tools for the treatment of Parkinson's disease (PD). Intrabodies make use of the specificity of antibodies to form complexes with intracellular proteins, and are already in clinical trials for treatment of cancers and AIDS. Recently, we published the first application of this powerful technology to a neurodegenerative disease, showing that human single-chain Fv intrabodies, selected from a phage display library, can counteract in situ Huntington aggregation in cellular models of Huntington's Disease (HD). By choosing an epitope adjacent to the abnormally folding expanded polyglutamine of the HD protein, we can alter the abnormal protein-protein interactions that characterize the mutant protein. Parkinson's Disease brains show formation of filamentous intracellular inclusions (Lewy bodies) as their most striking pathology. These appear to be composed of abnormally aggregated alpha-synuclein in both sporadic and hereditary forms of the disease. The alpha-synuclein protein data to date suggest several candidate target sequences that might prevent misfolding of synuclein, or be used to clear abnormal material in Parkinson's cells. We propose to select anti-synuclein single-chain Fv intrabodies against unique, defined domains and forms of a-syn, using human phage display libraries. These will then be tested for their capacity to reverse reported physiological defects in model systems. Models will include several that over-express wild-type alpha-synuclein to mimic sporadic P.D: transient transfections in cell lines and brain slice cultures, moving on to transgenic Drosophila and eventually transgenic mice. All of these studies will take advantage of the technologies and expertise coming out of the HD studies, which are actively and successfully ongoing in the lab. Long-term goals include administering these antibody reagents via gene therapy vectors, or as stable, multi-functional proteins.